The objective of this research is to develop a novel antibiotic targeting Gram positive bacteria, with an initial clinical indication of acute bacterial skin and skin structure infections (ABSSSI). The research will address the urgent problem of antibiotic resistant bacteria that are spreading around the world. The new antibiotic is being designed to inhibit a novel target, the bacterial methionyl- tRNA synthetase enzyme that is required for protein synthesis. The type of enzyme found in Gram positive bacteria is different from the comparable enzyme in the human cytoplasm, thus making it possible to design selective and safe inhibitors. In preliminary studies we have identified highly potent compounds against serious pathogens such as methicillin resistant Staphylococcus aureus (MRSA), vancomycin resistant Enterococcus faecium (VRE), Streptococcus species, and others. The minimum inhibitory concentrations (MICs) are below those of widely used drugs such as vancomycin and linezolid. The compounds have minimal toxicity on mammalian cells, are well tolerated in mice, are orally available and show efficacy in two mouse model of Staphylococcus infection comparable to linezolid. Specific Aim 1 will be to optimize the antibacterial, pharmacological, and safety properties of the compound in order to select a preclinical candidate. Approximately 150 new compounds will be synthesized and tested according to a screening cascade. The selected compound will pass safety tests such as Ames and hERG inhibition, and will demonstrate potent activity in the murine model of Staphylococcus infection. Specific Aim 2 will involve PK/ADME, in vivo efficacy testing in mice and in vitro toxicology studies to allow for identification of a lead candidate. This will be followed by definitive PK and exploratory toxicology studies in rats and evaluation of the lead candidate in additional efficacy studies (Aim 3). The scientific teams at University of Washington and TSRL have the combined expertise in chemistry, microbiology, pharmacology, and preclinical drug development to execute the proposed research plan. A successful project will bring forward a candidate drug in a novel antibiotic class to address the critical public health issue of bacterial drug-resistance.